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Etoposide sensitizes neuroblastoma cells expressing caspase 8 to TRAIL.

Kim HR, Lee MW, Kim DS, Jo HY, Lee SH, Chueh HW, Jung HL, Yoo KH, Sung KW, Koo HH - Cell Biol Int Rep (2010) (2012)

Bottom Line: Although TRAIL-R2 expression increased in IMR-32 cells in response to etoposide treatment, cell death was not increased by concurrent treatment with TRAIL compared with etoposide alone, because the cells lacked caspase 8 expression.Restoration of caspase 8 expression by exposure to IFNγ (interferon γ) sensitizes IMR-32 cells to TRAIL.Moreover, pretreatment with etoposide increased TRAIL-induced apoptosis in caspase 8 restored IMR-32 cells through activation of a caspase cascade that included caspases 8, 9 and 3.

View Article: PubMed Central - PubMed

Affiliation: Department of Pediatrics, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Korea.

ABSTRACT
TRAIL [TNF (tumour necrosis factor)-related apoptosis-inducing ligand] is a promising agent for clinical use since it kills a wide range of tumour cells without affecting normal cells. We provide evidence that pretreatment with etoposide significantly enhanced TRAIL-mediated apoptosis via up-regulation of DR5 (death receptor 5 or TRAIL-R2) expression in the caspase 8 expressing neuroblastoma cell line, SK-N-MC. In addition, sequential treatment with etoposide and TRAIL increased caspases 8, 9 and 3 activation, Mcl-1 cleavage and Bid truncation, which suggests that the ability of etoposide and TRAIL to induce apoptosis is mediated through activation of an intrinsic signalling pathway. Although TRAIL-R2 expression increased in IMR-32 cells in response to etoposide treatment, cell death was not increased by concurrent treatment with TRAIL compared with etoposide alone, because the cells lacked caspase 8 expression. Restoration of caspase 8 expression by exposure to IFNγ (interferon γ) sensitizes IMR-32 cells to TRAIL. Moreover, pretreatment with etoposide increased TRAIL-induced apoptosis in caspase 8 restored IMR-32 cells through activation of a caspase cascade that included caspases 8, 9 and 3. These results indicate that the etoposide-mediated sensitization of neuroblastoma cells to TRAIL is associated with an increase in TRAIL-R2 expression and requires caspase 8 expression. These observations support the potential use of a combination of etoposide and TRAIL in future clinical trials.

No MeSH data available.


Related in: MedlinePlus

Cells lacking caspase 8 expression are resistant to TRAIL-induced cell death(a) The expression of caspases (8, 9 and 3) and Bcl-2 family members (Mcl-1, Bid, Bax and Bcl-2) in IMR-32 and SK-N-MC cells after treatment with etoposide (500 ng/ml) and/or TRAIL (100 ng/ml) for 12 h. Equal loading was assessed by staining with anti-β-actin. (b) Cells were treated for 48 h with etoposide (Eto; 500 ng/ml) and/or TRAIL (T; 100 ng/ml) in the absence or presence of 10 μM zIETD-fmk (a caspase-8 inhibitor), 50 μM zLEHD-fmk (a caspase-9 inhibitor), or 50 μM zDEVD-fmk (a caspase-3 inhibitor). The results are presented as cell death as a percentage against vehicle-treated control (means±S.D. from three independent experiments).
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Figure 3: Cells lacking caspase 8 expression are resistant to TRAIL-induced cell death(a) The expression of caspases (8, 9 and 3) and Bcl-2 family members (Mcl-1, Bid, Bax and Bcl-2) in IMR-32 and SK-N-MC cells after treatment with etoposide (500 ng/ml) and/or TRAIL (100 ng/ml) for 12 h. Equal loading was assessed by staining with anti-β-actin. (b) Cells were treated for 48 h with etoposide (Eto; 500 ng/ml) and/or TRAIL (T; 100 ng/ml) in the absence or presence of 10 μM zIETD-fmk (a caspase-8 inhibitor), 50 μM zLEHD-fmk (a caspase-9 inhibitor), or 50 μM zDEVD-fmk (a caspase-3 inhibitor). The results are presented as cell death as a percentage against vehicle-treated control (means±S.D. from three independent experiments).

Mentions: Caspases 8, 9 and 3 activation and Mcl-1 cleavage were induced by TRAIL in SK-N-MC cells. Consecutive treatment with etoposide and TRAIL significantly increased the activation of caspases 8, 9 and 3, as well as Mcl-1 cleavage and Bid truncation, which all correlated with the increase in cell death (Figure 3a). Cell death decreased in the presence of caspases 8, 9 or 3 inhibitors in SK-N-MC cells (Figure 3b). In contrast, caspases 9 and 3 activation, Mcl-1 cleavage and Bid truncation were induced by etoposide alone in IMR-32 cells lacking caspase 8 expression. Moreover, etoposide-induced cell death decreased in the presence of caspase 9 or 3 inhibitors but was not affected by a caspase 8 inhibitor (Figure 3b). Consecutive treatment with etoposide and TRAIL did not increase activation of caspases, Mcl-1 cleavage, or Bid truncation compared with etoposide treatment alone. No substantial changes in Bcl-2 or Bax were detected under any of the treatment conditions in either cell line (Figure 3a).


Etoposide sensitizes neuroblastoma cells expressing caspase 8 to TRAIL.

Kim HR, Lee MW, Kim DS, Jo HY, Lee SH, Chueh HW, Jung HL, Yoo KH, Sung KW, Koo HH - Cell Biol Int Rep (2010) (2012)

Cells lacking caspase 8 expression are resistant to TRAIL-induced cell death(a) The expression of caspases (8, 9 and 3) and Bcl-2 family members (Mcl-1, Bid, Bax and Bcl-2) in IMR-32 and SK-N-MC cells after treatment with etoposide (500 ng/ml) and/or TRAIL (100 ng/ml) for 12 h. Equal loading was assessed by staining with anti-β-actin. (b) Cells were treated for 48 h with etoposide (Eto; 500 ng/ml) and/or TRAIL (T; 100 ng/ml) in the absence or presence of 10 μM zIETD-fmk (a caspase-8 inhibitor), 50 μM zLEHD-fmk (a caspase-9 inhibitor), or 50 μM zDEVD-fmk (a caspase-3 inhibitor). The results are presented as cell death as a percentage against vehicle-treated control (means±S.D. from three independent experiments).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3475444&req=5

Figure 3: Cells lacking caspase 8 expression are resistant to TRAIL-induced cell death(a) The expression of caspases (8, 9 and 3) and Bcl-2 family members (Mcl-1, Bid, Bax and Bcl-2) in IMR-32 and SK-N-MC cells after treatment with etoposide (500 ng/ml) and/or TRAIL (100 ng/ml) for 12 h. Equal loading was assessed by staining with anti-β-actin. (b) Cells were treated for 48 h with etoposide (Eto; 500 ng/ml) and/or TRAIL (T; 100 ng/ml) in the absence or presence of 10 μM zIETD-fmk (a caspase-8 inhibitor), 50 μM zLEHD-fmk (a caspase-9 inhibitor), or 50 μM zDEVD-fmk (a caspase-3 inhibitor). The results are presented as cell death as a percentage against vehicle-treated control (means±S.D. from three independent experiments).
Mentions: Caspases 8, 9 and 3 activation and Mcl-1 cleavage were induced by TRAIL in SK-N-MC cells. Consecutive treatment with etoposide and TRAIL significantly increased the activation of caspases 8, 9 and 3, as well as Mcl-1 cleavage and Bid truncation, which all correlated with the increase in cell death (Figure 3a). Cell death decreased in the presence of caspases 8, 9 or 3 inhibitors in SK-N-MC cells (Figure 3b). In contrast, caspases 9 and 3 activation, Mcl-1 cleavage and Bid truncation were induced by etoposide alone in IMR-32 cells lacking caspase 8 expression. Moreover, etoposide-induced cell death decreased in the presence of caspase 9 or 3 inhibitors but was not affected by a caspase 8 inhibitor (Figure 3b). Consecutive treatment with etoposide and TRAIL did not increase activation of caspases, Mcl-1 cleavage, or Bid truncation compared with etoposide treatment alone. No substantial changes in Bcl-2 or Bax were detected under any of the treatment conditions in either cell line (Figure 3a).

Bottom Line: Although TRAIL-R2 expression increased in IMR-32 cells in response to etoposide treatment, cell death was not increased by concurrent treatment with TRAIL compared with etoposide alone, because the cells lacked caspase 8 expression.Restoration of caspase 8 expression by exposure to IFNγ (interferon γ) sensitizes IMR-32 cells to TRAIL.Moreover, pretreatment with etoposide increased TRAIL-induced apoptosis in caspase 8 restored IMR-32 cells through activation of a caspase cascade that included caspases 8, 9 and 3.

View Article: PubMed Central - PubMed

Affiliation: Department of Pediatrics, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Korea.

ABSTRACT
TRAIL [TNF (tumour necrosis factor)-related apoptosis-inducing ligand] is a promising agent for clinical use since it kills a wide range of tumour cells without affecting normal cells. We provide evidence that pretreatment with etoposide significantly enhanced TRAIL-mediated apoptosis via up-regulation of DR5 (death receptor 5 or TRAIL-R2) expression in the caspase 8 expressing neuroblastoma cell line, SK-N-MC. In addition, sequential treatment with etoposide and TRAIL increased caspases 8, 9 and 3 activation, Mcl-1 cleavage and Bid truncation, which suggests that the ability of etoposide and TRAIL to induce apoptosis is mediated through activation of an intrinsic signalling pathway. Although TRAIL-R2 expression increased in IMR-32 cells in response to etoposide treatment, cell death was not increased by concurrent treatment with TRAIL compared with etoposide alone, because the cells lacked caspase 8 expression. Restoration of caspase 8 expression by exposure to IFNγ (interferon γ) sensitizes IMR-32 cells to TRAIL. Moreover, pretreatment with etoposide increased TRAIL-induced apoptosis in caspase 8 restored IMR-32 cells through activation of a caspase cascade that included caspases 8, 9 and 3. These results indicate that the etoposide-mediated sensitization of neuroblastoma cells to TRAIL is associated with an increase in TRAIL-R2 expression and requires caspase 8 expression. These observations support the potential use of a combination of etoposide and TRAIL in future clinical trials.

No MeSH data available.


Related in: MedlinePlus